Charged ion channel blockers and methods for use

ABSTRACT

The invention provides compounds of Formula (I), or pharmaceutically acceptable salts thereof. The compounds, compositions, methods and kits of the invention are particularly useful for the treatment of itch and other dermal conditions.

RELATED APPLICATION

This application is a continuation-in-part of U.S. application Ser. No.16/815,426 filed on Mar. 11, 2020. The entire contents of the aboveapplication are incorporated by reference herein.

TECHNICAL FIELD

The present invention relates generally to quaternary ammoniumcompounds, pharmaceutical compositions, and methods useful as selectiveinhibitors of pain, cough and itch sensing neurons (nociceptors, coughreceptors and pruriceptors) and in the treatment of neurogenicinflammation.

BACKGROUND OF THE INVENTION

The invention features compounds, compositions and methods for theselective inhibition of sensory neurons and the treatment of itch and/orneurogenic inflammation by targeting nociceptors with a small moleculedrug, while minimizing effects on non-nociceptive neurons or other typesof cells. According to the method of the invention, small, cationic drugmolecules gain access to the intracellular compartment of sensoryneurons via entry through large pore receptor/ion channels that arepresent in itch-sensing neurons but to a lesser extent or not at all inother types of neurons or in other types of tissue.

Local anesthetics such as lidocaine, articaine and pramocaine act byinhibiting voltage-dependent sodium channels in neurons. Theseanesthetics block sodium channels and thereby the excitability of allneurons, not just pain-sensing neurons (nociceptors). Thus, while thegoal of topical or regional anesthesia is to block transmission ofsignals in nociceptors to prevent pain, administration of localanesthetics also produces unwanted or deleterious effects such asgeneral numbness from block of low threshold pressure and touchreceptors, motor deficits and/or paralysis from block of motor axons andother complications from block of autonomic fibers. Local anestheticsare relatively hydrophobic molecules that gain access to their blockingsite on the sodium channel by diffusing through the cell membrane.Charged derivatives of these compounds, which are notmembrane-permeable, have no effect on neuronal sodium channels whenapplied to the external surface of the nerve membrane but can blocksodium channels if somehow introduced inside the cell, for example bydiffusion from a micropipette used for whole-cell electrophysiologicalrecording from isolated neurons. Pain-, cough-, and itch-sensing neuronsdiffer from other types of neurons in expressing (in most cases) theTRPV1 receptor/channel, which is activated by painful heat or bycapsaicin, the pungent ingredient in chili pepper. Other types ofchannels selectively expressed in various types of pain-sensing,cough-sensing and itch-sensing (pruriceptor) neurons include but are notlimited to TRPV2-4, TRPA1, TRPM8, ASIC and P2X(2/3) channels. It is wellestablished that some cationic small molecules such as QX-314 are ableto enter a cell via passage through activated large pore channels suchas TRPV1.

Neuropathic, inflammatory, and nociceptive pain differ in theiretiology, pathophysiology, diagnosis, and treatment. Nociceptive painoccurs in response to the activation of a specific subset of highthreshold peripheral sensory neurons, the nociceptors, by intense ornoxious stimuli. It is generally acute, self-limiting and serves aprotective biological function by acting as a warning of potential oron-going tissue damage. It is typically well-localized. Examples ofnociceptive pain include, but are not limited to, traumatic or surgicalpain, labor pain, sprains, bone fractures, burns, bumps, bruises,injections, dental procedures, skin biopsies, and obstructions.

Inflammatory pain is pain that occurs in the presence of tissue damageor inflammation including postoperative (i.e. pain associated with acuteperioperative pain resulting from inflammation caused by tissue trauma(e.g., surgical incision, dissection, burns) or direct nerve injury(e.g., nerve transection, stretching, or compression)), post-traumaticpain, arthritic pain (rheumatoid; or osteoarthritis (i.e. joint pain andstiffness due to gradual deterioration of the joint cartilage; riskfactors include aging, injury, and obesity; commonly affected joints arethe hand, wrist, neck, knee, hip, and spine), pain and pain associatedwith damage to joints, muscle, and tendons as in axial low back pain(i.e. a prevalent, painful condition affecting the lower portion of theback; common causes include muscle strain, spine fracture, bulging orruptured disc, and arthritis), severe nociceptive pain may transition toinflammatory pain if there is associated tissue injury.

Neuropathic pain is a common type of chronic, non-malignant pain, whichis the result of an injury or malfunction in the peripheral or centralnervous system and serves no protective biological function. It isestimated to affect more than 1.6 million people in the U.S. population.Neuropathic pain has many different etiologies, and may occur, forexample, due to trauma, surgery, herniation of an intervertebral disk,spinal cord injury, diabetes, infection with herpes zoster (shingles),HIV/AIDS, late-stage cancer, amputation (including mastectomy), carpaltunnel syndrome, chronic alcohol use, exposure to radiation, and as anunintended side-effect of neurotoxic treatment agents, such as certainanti-HIV and chemotherapeutic drugs. Peripheral neuropathy is caused bydamages to the peripheral nerves from injury, trauma, prolongedpressure, or inflammation causing numbness and pain in correspondingareas of the body.

Neuropathic pain is frequently described as “burning,” “electric,”“tingling,” or “shooting” in nature. It is often characterized bychronic dynamic allodynia (defined as pain resulting from a movingstimulus that does not ordinarily elicit a painful response, such aslight touch) and hyperalgesia (defined as an increased sensitivity to anormally painful stimulus) and may persist for months or years beyondthe apparent healing of any damaged tissues.

Pain may occur in patients with cancer, which may be due to multiplecauses; inflammation, compression, invasion, metastatic spread into boneor other tissues.

There are some conditions where pain occurs in the absence of a noxiousstimulus, tissue damage or a lesion to the nervous system, calleddysfunctional pain and these include but are not limited tofibromyalgia, tension type headache, and irritable bowel disorders.

Migraine is a headache associated with the activation of sensory fibersinnervating the meninges of the brain.

Itch (pruritus) is a dermatological condition that may be localized andgeneralized and can be associated with skin lesions (rash, atopiceczema, wheals). Itch accompanies many conditions including but notlimited to stress, anxiety, UV radiation from the sun, metabolic andendocrine disorders (e.g., liver or kidney disease, hyperthyroidism),cancers (e.g., lymphoma), reactions to drugs or food, parasitic andfungal infections, allergic reactions, diseases of the blood (e.g.,polycythemia vera), and dermatological conditions. Itch is mediated by asubset of small diameter primary sensory neurons, the pruriceptor, thatshare many features of nociceptor neurons, including but not limited toexpression of TRPV1 channels and other large pore channels (e.g.TRPV2-4, TRPA1, TRPM8, ASIC and P2X(2/3). Certain itch mediators—such aseicosanoids, histamine, bradykinin, ATP, and various neurotrophins haveendovanilloid functions. Topical capsaicin suppresses histamine-induceditch. Pruriceptors like nociceptors are therefore a suitable target forthis method of delivering ion channel blockers.

Cough is a defensive reflex designed to protect the airway from foreignbodies and to aid in the clearance of luminal debris. This reflex,however, can became aberrant in a number of diseases leading to anon-productive dry cough where hyper- or allo-tussive states exist.Hyper- and allo-tussive states are often chronic in nature lastinggreater than three months and can be manifested in many airway diseasesstates including asthma, COPD, asthma-COPD overlap syndrome (ACOS),interstitial pulmonary fibrosis (IPF) and lung cancer. In addition,inappropriate cough reflexes can be manifested acutely and chronicallyfollowing viral infection. Furthermore, chronic cough can be idiopathicin nature with unknown etiology.

Neurogenic inflammation is a mode of inflammation mediated by theefferent (motor) functions of sensory neurons, in which pro-inflammatorymediator molecules released in the periphery by pain-sensing neurons(nociceptors) activate a variety of inflammatory pathways in immunecells, and also act on the vascular system to alter blood flow andcapillary permeability.

Neurogenic inflammation contributes to peripheral inflammation elicitedby tissue injury, autoimmune disease, infection, allergy, exposure toirritants in a variety of tissues, and is thought to play an importantrole in the pathogenesis of numerous disorders (e.g. allergicinflammation, inflammatory bowel disease, interstitial cystitis, atopicdermatitis, asthma, conjunctivitis, arthritis, colitis, contactdermatitis, diabetes, eczema, cystitis, gastritis, migraine headache,psoriasis, rhinitis, rosacea, sunburn, pancreatitis, chronic cough,chronic rhinosinusistis, traumatic brain injury, polymicrobial sepsis,tendinopathies, chronic urticaria, rheumatic disease, acute lung injury,exposure to irritants, inhalation of irritants, pollutants, or chemicalwarfare agents). One way to reduce neurogenic inflammation is to blockexcitability in nociceptors, thereby preventing the activation ofnociceptor peripheral terminals and the release of pro-inflammatorymediators.

Despite the development of a variety of therapies for pain, itch, andneurogenic inflammation, there is a need for additional agents.

SUMMARY OF THE INVENTION

The present invention provides compounds represented by Formula (I) thatcan be used to treat or prevent itch, pain, cough, and neurogenicinflammation:

wherein:

Y⁻ is a pharmaceutically acceptable anion;

Wherein R₁, R₂, R₃, R₄, R₅, and R₆ are independently selected fromhydrogen, C1-C4 alkyl, C1-C4 heteroalkyl, aryl or heteroaryl

n is 0, 1, 2, 3, 4 and 5;

q is 0, 1, 2, 3, 4 or 5;

The invention includes the surprising finding that the compounds areparticularly active for the treatment of itch. The compounds have highpotency and superior topical localization, making the compoundssurprisingly suitable for dermal applications.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1A, 1B and 1C show that representative compounds significantlyreduce chloroquine-induced itch up to 8 hours following topicaladministration.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides compounds represented by Formula (I) asdescribed above, or pharmaceutically acceptable salts, stereoisomers,solvates, hydrates or combinations thereof. The invention also providescompositions comprising compounds having Formula (I) or apharmaceutically acceptable salts thereof, for example, a compositioncomprising an effective amount of a compound of Formula (I) or apharmaceutically acceptable salt thereof, and a pharmaceuticallyacceptable excipient. The compositions of the invention may furthercomprise compounds of the invention and a biologically active agent. Thecompositions can be formulated for oral, intravenous, intramuscular,rectal, cutaneous, subcutaneous, topical, transdermal, sublingual,nasal, inhalation, vaginal, intrathecal, epidural, or ocularadministration. However, preferred compositions can be formulated fortopical or dermal administration.

The invention further provides methods for treating itch, pruritis,psoriasis or atopic dermatitis, in a patient, including administering tothe patient a composition comprising a compound having Formula (I).

As used herein, the words “a” and “an” are meant to include one or moreunless otherwise specified.

By “biologically active” is meant that a molecule, including biologicalmolecules, such as nucleic acids, peptides, polypeptides, and proteins,exerts a biological, physical or chemical effect activity on a protein,enzyme, receptor, ligand, antigen, itself or other molecule. Forexample, a “biologically active” molecule may possess, e.g., enzymaticactivity, protein binding activity, or pharmacological activities.

Biologically active agents that can be used in the methods and kitsdescribed herein include, without limitation, TRPA1 receptor agonists,TRPV1-4 receptor agonists, ASIC agonists, TRPM8 agonists, P2X receptoragonists, NSAIDs, glucocorticoids, narcotics, anti-proliferative andimmune modulatory agents, an antibody or antibody fragment, anantibiotic, a polynucleotide, a polypeptide, a protein, an anti-canceragent, a growth factor, and a vaccine.

The term “itch” is used herein in the broadest sense and refers toitching and stinging sensations localized and generalized, acuteintermittent and persistent. The itch may be idiopathic, allergic,metabolic, infectious, drug-induced, due to liver, kidney disease, orcancer. “Pruritus” is severe itching.

By “inflammation” is meant any types of inflammation, such those causedby the immune system (immune-mediated inflammation) and by the nervoussystem (neurogenic inflammation), and any symptom of inflammation,including redness, heat, swelling, pain, and/or loss of function. Dermalinflammation is preferred.

By “neurogenic inflammation” is meant any type of inflammation mediatedor contributed to by neurons (e.g., nociceptors) or any other componentof the central or peripheral nervous system. Dermal neurogenicinflammation is preferred.

The term “pain” is used herein in the broadest sense and refers to alltypes of pain, including acute and chronic pain, such as nociceptivepain, e.g., somatic pain and visceral pain; inflammatory pain,dysfunctional pain, idiopathic pain, neuropathic pain, e.g., centrallygenerated pain and peripherally generated pain, migraine, and cancerpain. Dermal pain is preferred.

The term “nociceptive pain” is used to include all pain caused bynoxious stimuli that threaten to or actually injure body tissues,including, without limitation, by a cut, bruise, bone fracture, crushinjury, burn, and the like. Pain receptors for tissue injury(nociceptors) are located mostly in the skin, musculoskeletal system, orinternal organs, preferably the skin or dermis.

The term “somatic pain” is used to refer to pain arising from bone,joint, muscle, skin, or connective tissue, preferably the skin andtissues adjacent to the skin. This type of pain is typically welllocalized.

The term “visceral pain” is used herein to refer to pain arising fromvisceral organs, such as the respiratory, gastrointestinal tract andpancreas, the urinary tract and reproductive organs. Visceral painincludes pain caused by tumor involvement of the organ capsule. Anothertype of visceral pain, which is typically caused by obstruction ofhollow viscus, is characterized by intermittent cramping and poorlylocalized pain. Visceral pain may be associated with inflammation as incystitis or reflux esophagitis.

The term “inflammatory pain” includes pain associates with activeinflammation that may be caused by trauma, surgery, infection andautoimmune diseases.

The term “neuropathic pain” is used herein to refer to pain originatingfrom abnormal processing of sensory input by the peripheral or centralnervous system consequent on a lesion to these systems.

The term “procedural pain” refers to pain arising from a medical, dentalor surgical procedure wherein the procedure is usually planned orassociated with acute trauma.

By “patient” is meant any animal. In one embodiment, the patient is ahuman. Other animals that can be treated using the methods,compositions, and kits of the invention include but are not limited tonon-human primates (e.g., monkeys, gorillas, chimpanzees), domesticatedanimals (e.g., horses, pigs, goats, rabbits, sheep, cattle, llamas), andcompanion animals (e.g., guinea pigs, rats, mice, lizards, snakes, dogs,cats, fish, hamsters, and birds).

Compounds useful in the invention include, but are not limited to, thosedescribed herein in any of their pharmaceutically acceptable forms,including isomers such as diastereomers and enantiomers, salts, esters,amides, thioesters, solvates, and polymorphs thereof, as well as racemicmixtures and pure isomers of the compounds described herein.

The term “pharmaceutically acceptable anion” as used herein, refers tothe conjugate base of a pharmaceutically acceptable acid. Such acids aredescribed in Stahl, P. H. and Wermuth, C. G. (eds.), Handbook ofPharmaceutical Salts: Properties, Selection and Use, Wiley VCH (2008).Pharmaceutically acceptable acids include, but are not limited to,acetic acid, dichloroacetic acid, adipic acid, alginic acid, L-ascorbicacid, L-aspartic acid, benzenesulfonic acid, 4-acetamidobenzoic acid,benzoic acid, p-bromophenylsulfonic acid, (+)-camphoric acid,(+)-camphor-10-sulfonic acid, capric acid, caproic acid, caprylic acid,carbonic acid, cinnamic acid, cyclamic acid, dodecylsulfuric acid,ethane-1,2-disulfonic acid, ethanesulfonic acid, 2-hydroxyethanesulfonicacid, sulfuric acid, boric acid, citric acid, formic acid, fumaric acid,galactaric acid, gentisic acid, D-glucoheptonic acid, D-gluconic acid,D-glucuronic acid, glutamic acid, glutaric acid, 2-oxoglutaric acid,glycerophosphoric acid, glycolic acid, hippuric acid, hydrochloric acid,hydrobromic acid, hydroiodic acid, isobutyric acid, DL-lactic acid,lactobionic acid, lauric acid, maleic acid, (−)-L-malic acid, malonicacid, DL-mandelic acid, methanesulfonic acid, naphthalene-1,5-disulfonicacid, naphthalene-2-sulfonic acid, 1-hydroxy-2-naphthoic acid, nicotinicacid, nitric acid, oleic acid, orotic acid, oxalic acid, palmitic acid,pamoic acid, phosphoric acid, propionic acid, (−)-L-pyroglutamic acid,salicyclic acid, 4-aminosalicyclic acid, sebacic acid, stearic acid,succinic acid, (+)-L-tartaric acid, thiocyanic acid, p-toluenesulfonicacid, and undecylenic acid. Pharmaceutically acceptable anions includethe conjugate base of any the acids set forth above.

The term “pharmaceutically acceptable salt” represents those salts whichare, within the scope of sound medical judgment, suitable for use incontact with the tissues of humans and lower animals without unduetoxicity, irritation, allergic response and the like, and arecommensurate with a reasonable benefit/risk ratio. The salts can beprepared in situ during the final isolation and purification of thecompounds of the invention, or separately by reacting the free basefunction with a suitable organic acid. Representative acid additionsalts include but are not limited to acetate, adipate, alginate,ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate,butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate,digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptonate,glycerophosphate, hemisulfate, heptonate, hexanoate, hydrobromide,hydrochloride, hydroiodide, 2-hydroxy-ethanesulfonate, isethionate,lactobionate, lactate, laurate, lauryl sulfate, malate, maleate,malonate, mesylate, methanesulfonate, 2-naphthalenesulfonate,nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate,persulfate, 3-phenylpropionate, phosphate, picrate, pivalate,propionate, stearate, succinate, sulfate, tartrate, thiocyanate,toluenesulfonate, undecanoate, valerate salts, and the like.

By “charged moiety” is meant a moiety which gains a proton atphysiological pH thereby becoming positively charged (e.g., ammonium,guanidinium, or amidinium) or a moiety that includes a net formalpositive charge without protonation (e.g., quaternary ammonium). Thecharged moiety may be either permanently charged or transiently charged.

By “therapeutically effective amount” or “effective amount” means anamount sufficient to produce a desired result, for example, thereduction or elimination of pain, cough, itch, or neurogenicinflammation in a patient (e.g., a human) suffering from a condition,disease, or illness that is caused wholly or in part by neurogenicinflammation (e.g. asthma, arthritis, colitis, contact dermatitis,diabetes, eczema, cystitis, chronic refractory cough, post-viral cough,gastritis, migraine headache, psoriasis, rhinitis, rosacea, or sunburn).

“Solvates” means solvent addition forms that contain eitherstoichiometric or nonstoichiometric amounts of solvent.

The compounds of the present invention, including salts of thecompounds, can exist in unsolvated forms as well as solvated forms,including hydrated forms and unhydrated forms. In general, the solvatedforms are equivalent to unsolvated forms and are encompassed within thescope of the present invention. Nonlimiting examples of hydrates includemonohydrates, dihydrates, hemihydrates, etc. In certain aspects, thecompound is a hemihydrate. Nonlimiting examples of solvates includeethanol solvates, acetone solvates, etc.

The compounds of the invention may exist in multiple crystalline oramorphous forms. In general, all physical forms are equivalent for usescontemplated by the present invention and are intended to be within thescope of the invention.

Compounds that can be used in the compositions, kits, and methods of theinvention include compounds having Formula (I):

wherein:

Y⁻ is a pharmaceutically acceptable anion;

Wherein R₁, R₂, R₃, R₄, R₅, and R₆ are independently selected fromhydrogen, C1-C4 alkyl, C1-C4 heteroalkyl, aryl or heteroaryl

n is 0, 1, 2, 3, 4 and 5;

q is 0, 1, 2, 3, 4 or 5;

Preferably, Y⁻ is a halide anion, a carboxylate, or a sulfonate. Y⁻ can,for example, be a halide ion, a substituted or unsubstitutedalkylsulfonate, a substituted or unsubstituted arylsulfonate, asubstituted or unsubstituted alkyl or aliphatic carboxylate, asubstituted or unsubstituted aryl carboxylate, or a substituted orunsubstituted heterocyclyl carboxylate.

In certain embodiments, Y⁻ is selected from the group consisting oftrifluoroacetate, sulfate, phosphate, acetate, fumarate, formate,carbonate, maleate, citrate, pyruvate, succinate, oxalate, a sulfonate,(for example, methanesulfonate, trifluoromethanesulfonate,toluenesulfonate such as p-toluenesulfonate, benzenesulfonate,ethanesulfonate, camphorsulfonate, 2-mesitylenesulfonate, ornaphthalenesulfonate such as 2-naphthalenesulfonate), bisulfate,malonate, xinafoate, ascorbate, oleate, nicotinate, saccharinate,adipate, formate, glycolate, L-lactate, D-lactate, aspartate, malate,L-tartrate, D-tartrate, stearate, 2-furoate, 3-furoate, napadisylate(naphthalene-1,5-disulfonate or naphthalene-1-(sulfonicacid)-5-sulfonate), edisylate (ethane-1,2-disulfonate orethane-1-(sulfonic acid)-2-sulfonate), isethionate(2-hydroxyethylsulfonate), D-mandelate, L-mandelate, propionate,tartarate, phthalate, hydrochlorate, hydrobromate, and nitrate.

In one embodiment, Y⁻ is halide anion. In a preferred embodiment, Y⁻ isselected from the halide ions bromide, chloride, or iodide.

As will be understood, when q is 0, 1, 2, 3, 4 or 5, the N⁺-containingring of Formula (I) is:

respectively. Preferably, q is 3. Another preferred embodiment, q is 2.Alternatively, q is 0, 1, 4 or 5.

In Formula I, R₁, R₂, R₃, R₄, R₅, and/or R₆ can each be hydrogen (e.g.,forming a straight chain alkylenyl, such as ethylenyl, propylenyl,butylenyl or pentylenyl). In preferred compounds, each of R₅ and R₆ arehydrogen. In other compounds, the R₁ and/or R₂ are both hydrogen. Inother compounds, the R₃ and R₄ are both hydrogen. Alternatively, R₁ ismethyl or ethyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen. Alternatively,R₃ is methyl or ethyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen.

For example, q is 0 and R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

q is 0 and R₁ is methyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

q is 0 and R₃ is methyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen; or

q is 1 and R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

q is 1 and R₁ is methyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

q is 1 and R₃ is methyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen; or

q is 2 and R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

q is 2 and R₁ is methyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

q is 2 and R₃ is methyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen; or

q is 3 and R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

q is 3 and R₁ is methyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

q is 3 and R₃ is methyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen.

For example, n is 0 and R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

n is 0 and R₁ is methyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

n is 0 and R₃ is methyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen; or

n is 1 and R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

n is 1 and R₁ is methyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

n is 1 and R₃ is methyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen; or

n is 2 and R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

n is 2 and R₁ is methyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

n is 2 and R₃ is methyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen; or

n is 3 and R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

n is 3 and R₁ is methyl and R₂, R₃, R₄, R₅, and R₆ are hydrogen; or

n is 3 and R₃ is methyl and R₁, R₂, R₄, R₅, and R₆ are hydrogen.

Each preferred group stated above can be taken in combination with one,any or all other preferred groups.

Compositions of the invention where R is methyl or ethyl can compriseracemic mixtures, pure enantiomers, or an excess of one enantiomer overthe other. For example, a composition can comprise an enantiomericexcess of at least 5, 10, 20, 30, 40, 50, 60, 70, 80 or 90%. In oneembodiment, the enantiomeric excess is at least 95%.

The compounds of the invention include all enantiomers which may bedefined, in terms of absolute stereochemistry, as (R)- or (S)-, as wellas their racemic and optically pure forms, and is not limited to thosedescribed herein in any of their pharmaceutically acceptable forms,including enantiomers, salts, solvates, polymorphs, solvatomorphs,hydrates, anhydrous and other crystalline forms and combinationsthereof.

Preferably, a pharmaceutical composition comprises a compound of theinvention as an R enantiomer in substantially pure form; or, apharmaceutical composition comprises a compound of the invention as an Senantiomer in substantially pure form; or, a pharmaceutical compositioncomprises a compound of the invention as enantiomeric mixtures whichcontain an excess of the R enantiomer or an excess of the S enantiomer.It is particularly preferred that the pharmaceutical compositioncontains a compound of the invention which is a substantially pureoptical isomer. For the avoidance of doubt, a compound of the inventioncan, if desired, be used in the form of solvates.

Compounds having Formula (I) can be prepared using methods analogous tothat described in the Examples and the following synthetic schemes:

Additional Biologically Active Agents and Exogenous Large Pore ChannelAgonists

As described above, the compound or composition of the invention can beadministered with a biologically active agent. For example, one or moreadditional biologically active agents, including those typically used totreat neurogenic inflammation, may be used in combination with acompound or composition of the invention described herein. Thebiologically active agents include, but are not limited to, TRPA1receptor agonists, TRPV1-4 receptor agonists, TRPM8 agonists, ASICagonists, P2X receptor agonists, acetaminophen, NSAIDs, glucocorticoids,narcotics, tricyclic antidepressants, amine transporter inhibitors,anticonvulsants, anti-proliferative and immune modulatory agents, anantibody or antibody fragment, an antibiotic, a polynucleotide, apolypeptide, a protein, an anti-cancer agent, a growth factor, and avaccine.

TRPV1 agonists that can be employed in the methods, kits andcompositions of the invention include, but are not limited to, any thatactivates TRPV1 receptors on nociceptors and allows for entry of atleast one inhibitor of voltage-gated ion channels (for example, acompound of the invention). A suitable TRPV1 agonist is capsaicin oranother capsaicinoids, which are members of the vanilloid family ofmolecules. Naturally occurring capsaicinoids are capsaicin itself,dihydrocapsaicin, nordihydrocapsaicin, homodihydrocapsaicin,homocapsaicin, and nonivamide. Other suitable capsaicinoids andcapsaicinoid analogs and derivatives for use in the compositions andmethods of the present invention include naturally occurring andsynthetic capsaicin derivatives and analogs including, e.g., vanilloids(e.g., N-vanillyl-alkanedienamides, N-vanillyl-alkanedienyls, andN-vanillyl-cis-monounsaturated alkenamides), capsiate, dihydrocapsiate,nordihydrocapsiate and other capsinoids, capsiconiate,dihydrocapsiconiate and other coniferyl esters, capsiconinoid,resiniferatoxin, tinyatoxin, civamide, N-phenylmethylalkenamidecapsaicin derivatives, olvanil,N-[(4-(2-aminoethoxy)-3-methoxyphenyl)methyl]-9Z-octa-decanamide,N-oleyl-homovanillamide, triprenyl phenols (e.g., scutigeral),gingerols, piperines, shogaols, guaiacol, eugenol, zingerone, nuvanil,NE-19550, NE-21610, and NE-28345. Additional capsaicinoids, theirstructures, and methods of their manufacture are described in U.S. Pat.Nos. 7,446,226 and 7,429,673, which are hereby incorporated byreference.

Additional suitable TRPV1 agonists include but are not limited toeugenol, arvanil (N-arachidonoylvanillamine), anandamide,2-aminoethoxydiphenyl borate (2APB), AM404, resiniferatoxin, phorbol12-phenylacetate 13-acetate 20-homovanillate (PPAHV), olvanil (NE19550), OLDA (N-oleoyldopamine), N-arachidonyldopamine (NADA),6′-iodoresiniferatoxin (6′-IRTX), C18 N-acylethanolamines, lipoxygenasederivatives such as 12-hydroperoxyeicosatetraenoic acid, inhibitorcysteine knot (ICK) peptides (vanillotoxins), piperine, MSK195(N-[2-(3,4-dimethylbenzyl)-3-(pivaloyloxy)propyl]-2-[4-(2-aminoethoxy)-3-methoxyphenyl]acetamide),JYL79(N-[2-(3,4-dimethylbenzyl)-3-(pivaloyloxy)propyl]-N′-(4-hydroxy-3-methoxybenzyl)thiourea),hydroxy-alpha-sanshool, 2-aminoethoxydiphenyl borate, 10-shogaol,oleylgingerol, oleylshogaol, and SU200(N-(4-tert-butylbenzyl)-N′-(4-hydroxy-3-methoxybenzyl)thiourea). Stillother TRPV1 agonists include amylocaine, articaine, benzocaine,bupivacaine, carbocaine, carticaine, chloroprocaine, cyclomethycaine,dibucaine (cinchocaine), dimethocaine (larocaine), etidocaine,hexylcaine, levobupivacaine, lidocaine, mepivacaine, meprylcaine(oracaine), metabutoxycaine, piperocaine, prilocaine, procaine(novacaine), proparacaine, propoxycaine, risocaine, ropivacaine,tetracaine (amethocaine), and trimecaine.

Suitable TRPV2-4 agonists include, but are not limited to, are 2-APB,cannabinol, diphenylboronic anhydride, insulin-like growth factor 1,lysophosphatidylcholine, lysophosphatidylinositol, probenecid,Δ9-tetrahydrocannabinol, vanillin, eugenol, cinnamaldehyde, camphor,carvacrol, thymol, citral, farnesyl diphosphate, tetrahydrocannabivarin,incensole acetate, diphenylboronic anhydride, 6-tert-butyl-m-cresol,dihydrocarveocarveol, borneol, (−)-menthol, GSK1016790A, 4α-PDH,5,6-epoxyeicosatrienoic acid, 4α-PDD, bisandrographolide, citric acid,phorbol 12-myristate 13-acetate and RN1747.

Suitable TRPM8 agonists include, but are not limited to, are menthol,icilin, eucalyptus, linalool, geraniol, hydroxy-citronellal, WS-3,WS-23, Frescolat MGA, Frescolat ML, PMD 38, CPS125, Coolact P, M8-Ag,AITC, cryosim-3, AX-8 and Cooling Agent 10.

Suitable ASIC agonists include, but are not limited to,chlorophenylguanidine hydrochloride, GMQ hydrochloride,tetrahydropapaveroline (THP), reticulin, polyamine agmatine,lysophosphatidylcholine, arachidonic acid and neuropeptide SF.

Other biologically active agents which can be employed in the methods,compositions, and kits of the invention include any that activates TRPA1receptors on nociceptors or pruriceptors and allows for entry of atleast one inhibitor of voltage-gated ion channels. Suitable TRPA1agonists include but are not limited to cinnamaldehyde,allyl-isothiocynanate (mustard oil), diallyl disulfide, icilin, cinnamonoil, wintergreen oil, clove oil, acrolein, hydroxy-alpha-sanshool,2-aminoethoxydiphenyl borate, 4-hydroxynonenal, methylp-hydroxybenzoate, and 3′-carbamoylbiphenyl-3-yl cyclohexylcarbamate(URB597).

P2X agonists that can be employed in the methods, compositions, and kitsof the invention include any that activates P2X receptors on nociceptorsor pruriceptors and allows for entry of at least one inhibitor ofvoltage-gated ion channels. Suitable P2X agonists include but are notlimited to ATP, α,β-methylene ATP, 2-methylthio-ATP, 2′ and3′-O-(4-benzoylbenzoyl)-ATP, and ATP5′-O-(3-thiotriphosphate).

Other biologically active agents that can be used in combination withthe compounds of the invention include NSAIDs, glucocorticoids,narcotics, tricyclic antidepressants, amine transporter inhibitors,anticonvulsants, anti-proliferative and immune modulatory agents, anantibody or antibody fragment, an antibiotic, a polynucleotide, apolypeptide, a protein, an anti-cancer agent, a growth factor, and avaccine.

Non-steroidal anti-inflammatory drugs (NSAIDs) that can be administeredto a patient (e.g., a human) suffering from neurogenic inflammation incombination with a composition of the invention include, but are notlimited to, acetylsalicylic acid, amoxiprin, benorylate, benorilate,choline magnesium salicylate, diflunisal, ethenzamide, faislamine,methyl salicylate, magnesium salicylate, salicyl salicylate,salicylamide, diclofenac, aceclofenac, acemethacin, alclofenac,bromfenac, etodolac, indometacin, nabumetone, oxametacin, proglumetacin,sulindac, tolmetin, ibuprofen, alminoprofen, benoxaprofen, carprofen,dexibuprofen, dexketoprofen, fenbufen, fenoprofen, flunoxaprofen,flurbiprofen, ibuproxam, indoprofen, ketoprofen, ketorolac, loxoprofen,naproxen, oxaprozin, pirprofen, suprofen, tiaprofenic acid, mefenamicacid, flufenamic acid, meclofenamic acid, tolfenamic acid,phenylbutazone, ampyrone, azapropazone, clofezone, kebuzone, metamizole,mofebutazone, oxyphenbutazone, phenazone, sulfinpyrazone, piroxicam,droxicam, lornoxicam, meloxicam, tenoxicam, and the COX-2 inhibitorscelecoxib, etoricoxib, lumiracoxib, parecoxib, rofecoxib, valdecoxib,and pharmaceutically acceptable salts thereof.

Glucocorticoids that can be administered to a patient (e.g., a human)suffering from neurogenic inflammation in combination with a compositionof the invention include, but are not limited to, hydrocortisone,cortisone acetate, prednisone, prednisolone, methylprednisolone,dexamethasone, betamethasone, triamcinolone, beclometasone,fludrocortisone acetate, deoxycorticosterone acetate, aldosterone, andpharmaceutically acceptable salts thereof.

Narcotics that can be administered to a patient (e.g., a human)suffering from neurogenic inflammation in combination with a compositionof the invention include, but are not limited, to tramadol, hydrocodone,oxycodone, morphine, and pharmaceutically acceptable salts thereof.

Antiproliferative and immune modulatory agents that can be administeredto a patient (e.g., a human) suffering from neurogenic inflammation incombination with a composition of the invention include, but are notlimited to, alkylating agents, platinum agents, antimetabolites,topoisomerase inhibitors, dihydrofolate reductase inhibitors, antitumorantibiotics, antimitotic agents, aromatase inhibitors, thymidylatesynthase inhibitors, DNA antagonists, farnesyltransferase inhibitors,pump inhibitors, histone acetyltransferase inhibitors, metalloproteinaseinhibitors, ribonucleoside reductase inhibitors, TNF-alpha agonists,TNF-alpha antagonists or scavengers, interleukin 1 (IL-1) antagonists orscavengers, endothelin A receptor antagonists, retinoic acid receptoragonists, hormonal agents, antihormonal agents, photodynamic agents, andtyrosine kinase inhibitors.

The biologically active agents can be administered prior to, concurrentwith, or following administration of a composition of the invention,using any formulation, dosing, or administration known in the art thatis therapeutically effective.

Formulation of Compositions

The administration of the compounds of the invention may be by anysuitable means that results in the reduction of perceived pain or itchsensation at the target region. The compounds of the invention may becontained in any appropriate amount in any suitable carrier substance,and are generally present in amounts totaling 1-99% by weight of thetotal weight of the composition. The composition may be provided in adosage form that is suitable for oral, parenteral (e.g., intravenous,intramuscular), rectal, cutaneous, subcutaneous, topical, transdermal,sublingual, nasal, vaginal, intrathecal, epidural, or ocularadministration, or by injection, inhalation, or direct contact with thenasal or oral mucosa. Topical or dermal administration is preferred.

Thus, the composition may be in the form of, e.g., suspensions,emulsions, solutions, gels including hydrogels, pastes, ointments,creams, plasters, sprays, aerosols, drenches, osmotic delivery devices,suppositories, enemas, injectables, implants, tablets, capsules, pills,powders, granulates. The compositions may be formulated according toconventional pharmaceutical practice (see, e.g., Remington: The Scienceand Practice of Pharmacy, 22nd edition, 2013, ed. L. V. Allen,Pharmaceutical Press, Philadelphia, and Encyclopedia of PharmaceuticalTechnology, 4^(th) Edition, ed. J. Swarbrick, 2013, CRC Press, NewYork).

Each compound may be formulated in a variety of ways that are known inthe art. For example, a compound of the invention and a biologicallyactive agent as defined herein may be formulated together or separately.Desirably, a compound of the invention and a biologically active agentare formulated together for their simultaneous or near simultaneousadministration. In another embodiment, two or more biologically activeagents may be formulated together with a compound of the invention, orseparately. Other examples include, but are not limited to, two or morecompounds of the invention formulated together, wherein the compoundsare formulated together with or without one or more biologically activeagents.

The individually or separately formulated agents can be packagedtogether as a kit. Non-limiting examples include but are not limited tokits that contain, e.g., two pills, a pill and a powder, a suppositoryand a liquid in a vial, two topical creams, etc. The kit can includeoptional components that aid in the administration of the unit dose topatients, such as vials for reconstituting powder forms, syringes forinjection, customized IV delivery systems, inhalers, etc. Additionally,the unit dose kit can contain instructions for preparation andadministration of the compositions.

The kit may be manufactured as a single use unit dose for one patient,multiple uses for a particular patient (at a constant dose or in whichthe individual compounds may vary in potency as therapy progresses); orthe kit may contain multiple doses suitable for administration tomultiple patients (“bulk packaging”). The kit components may beassembled in cartons, blister packs, bottles, tubes, and the like.

Topical Formulations

The compositions of the invention, alone or in combination with one ormore of the biologically active agents described herein, can also beadapted for topical use with a topical vehicle containing from between0.0001% and 25% (w/w) or more of active ingredient(s).

In a preferred combination, the active ingredients are preferably eachfrom between 0.0001% to 10% (w/w), more preferably from between 0.0005%to 4% (w/w) active agent. The topical formulation, including but notlimited to a cream, gel, or ointment, can be applied one to four timesdaily, or as needed. Performing the methods described herein, thetopical vehicle containing the composition of the invention, or acombination therapy containing a composition of the invention ispreferably applied to the site of inflammation on the patient. Forexample, a cream may be applied to the hands of a patient suffering fromitch, pruritis, psoriasis, or atopic dermatitis.

The compositions can be formulated using any dermatologically acceptablecarrier. Exemplary carriers include a solid carrier, such as alumina,clay, microcrystalline cellulose, silica, or talc; and/or a liquidcarrier, such as an alcohol, a glycol, or a water-alcohol/glycol blend.The therapeutic agents may also be administered in liposomalformulations that allow therapeutic agents to enter the skin. Suchliposomal formulations are described in U.S. Pat. Nos. 5,169,637;5,000,958; 5,049,388; 4,975,282; 5,194,266; 5,023,087; 5,688,525;5,874,104; 5,409,704; 5,552,155; 5,356,633; 5,032,582; 4,994,213;8,822,537, and PCT Publication No. WO 96/40061. Examples of otherappropriate vehicles are described in U.S. Pat. Nos. 4,877,805,8,822,537, and EP Publication No. 0586106A1. Suitable vehicles of theinvention may also include mineral oil, petrolatum, polydecene, stearicacid, isopropyl myristate, polyoxyl 40 stearate, stearyl alcohol, orvegetable oil.

The composition can further include a skin penetrating enhancer, such asthose described in “Percutaneous Penetration enhancers”, (eds. Smith E Wand Maibach H I. CRC Press 1995). Exemplary skin penetrating enhancersinclude alkyl (N,N-disubstituted amino alkanoate) esters, such asdodecyl 2-(N,N dimethylamino) propionate (DDAIP), which is described inU.S. Pat. Nos. 6,083,996 and 6,118,020, which are both incorporatedherein by reference; a water-dispersible acid polymer, such as apolyacrylic acid polymer, a carbomer (e.g., Carbopol™ or Carbopol 940P™,available from B. F. Goodrich Company (Akron, Ohio)), copolymers ofpolyacrylic acid (e.g., Pemulen™ from B. F. Goodrich Company orPolycarbophil™ from A. H. Robbins, Richmond, Va.; a polysaccharide gum,such as agar gum, alginate, carrageenan gum, ghatti gum, karaya gum,kadaya gum, rhamsan gum, xanthan gum, and galactomannan gum (e.g., guargum, carob gum, and locust bean gum), as well as other gums known in theart (see for instance, Industrial Gums: Polysaccharides & TheirDerivatives, Whistler R. L., BeMiller J. N. (eds.), 3rd Ed. AcademicPress (1992) and Davidson, R. L., Handbook of Water-Soluble Gums &Resins, McGraw-Hill, Inc., N.Y. (1980)); or combinations thereof.

Other suitable polymeric skin penetrating enhancers are cellulosederivatives, such as ethyl cellulose, methyl cellulose, hydroxypropylcellulose. Additionally, known transdermal penetrating enhancers canalso be added, if desired. Illustrative are dimethyl sulfoxide (DMSO)and dimethyl acetamide (DMA), 2-pyrrolidone, N,N-diethyl-m-toluamide(DEET), 1-dodecylazacycloheptane-2-one (Azone™, a registered trademarkof Nelson Research), N,N-dimethylformamide, N-methyl-2-pyrrolidone,calcium thioglycolate and other enhancers such as dioxolanes, cyclicketones, and their derivatives and so on.

Also illustrative are a group of biodegradable absorption enhancerswhich are alkyl N,N-2-(disubstituted amino) alkanoates as described inU.S. Pat. Nos. 4,980,378 and 5,082,866, which are both incorporatedherein by reference, including: tetradecyl (N,N-dimethylamino) acetate,dodecyl (N,N-dimethylamino) acetate, decyl (N,N-dimethylamino) acetate,octyl (N,N-dimethylamino) acetate, and dodecyl (N,N-diethylamino)acetate.

Particularly preferred skin penetrating enhancers include isopropylmyristate; isopropyl palmitate; dimethyl sulfoxide; decyl methylsulfoxide; dimethylalanine amide of a medium chain fatty acid; dodecyl2-(N,N-dimethylamino) propionate or salts thereof, such as its organic(e.g., hydrochloric, hydrobromic, sulfuric, phosphoric, and nitric acidaddition salts) and inorganic salts (e.g., acetic, benzoic, salicylic,glycolic, succinic, nicotinic, tartaric, maleic, malic, pamoic,methanesulfonic, cyclohexanesulfamic, picric, and lactic acid additionsalts), as described in U.S. Pat. No. 6,118,020; and alkyl2-(N,N-disubstituted amino)-alkanoates, as described in U.S. Pat. Nos.4,980,378 and 5,082,866.

The skin penetrating enhancer in this composition by weight would be inthe range of 0.5% to 10% (w/w). The most preferred range would bebetween 1.0% and 5% (w/w). In another embodiment, the skin penetratingenhancer comprises between 0.5%-1%, 1%-2%, 2%-3%, 3%-4%, or 4%-5%, (w/w)of the composition.

The compositions can be provided in any useful form. For example, thecompositions of the invention may be formulated as solutions, emulsions(including microemulsions), suspensions, creams, ointments, foams,lotions, gels, powders, or other typical solid, semi-solid, or liquidcompositions (e.g., topical sprays) used for application to the skin orother tissues where the compositions may be used. Such compositions maycontain other ingredients typically used in such products, such ascolorants, fragrances, thickeners (e.g., xanthan gum, a fatty acid, afatty acid salt or ester, a fatty alcohol, a modified cellulose, amodified mineral material, Krisgel 100™, or a synthetic polymer),antimicrobials, solvents, surfactants, detergents, gelling agents,antioxidants, fillers, dyestuffs, viscosity-controlling agents,preservatives, humectants, emollients (e.g., natural or synthetic oils,hydrocarbon oils, waxes, or silicones), hydration agents, chelatingagents, demulcents, solubilizing excipients, adjuvants, dispersants,skin penetrating enhancers, plasticizing agents, preservatives,stabilizers, demulsifiers, wetting agents, sunscreens, emulsifiers,moisturizers, astringents, deodorants, and optionally includinganesthetics, anti-itch actives, botanical extracts, conditioning agents,darkening or lightening agents, glitter, humectants, mica, minerals,polyphenols, silicones or derivatives thereof, sunblocks, vitamins, andphytomedicinals.

The compositions can also include other like ingredients to provideadditional benefits and improve the feel and/or appearance of thetopical formulation. Specific classes of additives commonly use in theseformulations include: isopropyl myristate, sorbic acid NF powder,polyethylene glycol, phosphatidylcholine (including mixtures ofphosphatidylcholine, such as phospholipon G), Krisgel 100™ distilledwater, sodium hydroxide, decyl methyl sulfoxide (as a skin penetratingenhancer), menthol crystals, lavender oil, butylated hydroxytoluene,ethyl diglycol reagent, and 95% percent (190 proof) ethanol.

Controlled Release Formulations

Each compound of the invention, alone or in combination with one or moreof the biologically active agents as described herein, can be formulatedfor controlled release (e.g., sustained or measured) administration, asdescribed in U.S. Patent Application Publication Nos. 2003/0152637 and2005/0025765, each incorporated herein by reference. For example, acompound of the invention, alone or in combination with one or more ofthe biologically active agents as described herein, can be incorporatedinto a patch, capsule or tablet that is administered to the patient.

Any pharmaceutically acceptable vehicle or formulation suitable forlocal application and/or injection into a site to be treated (e.g., apainful surgical incision, wound, or joint), that is able to provide asustained release of compound of the invention, alone or in combinationwith one or more of the biologically active agents as described herein,may be employed to provide for prolonged elimination or alleviation ofinflammation, as needed. Controlled release formulations known in theart include specially coated pellets, polymer formulations or matricesfor surgical insertion or as sustained release microparticles, e.g.,microspheres or microcapsules, for implantation, insertion, infusion orinjection, wherein the slow release of the active medicament is broughtabout through sustained or controlled diffusion out of the matrix and/orselective breakdown of the coating of the preparation or selectivebreakdown of a polymer matrix. Other formulations or vehicles forcontrolled, sustained, or immediate delivery of an agent to a preferredlocalized site in a patient include, e.g., suspensions, emulsions, gels,liposomes and any other suitable art known delivery vehicle orformulation acceptable for topical, transdermal, subcutaneous orintramuscular administration.

A wide variety of biocompatible materials may be utilized as acontrolled release carrier to provide the controlled release of acompound of the invention, alone or in combination with one or morebiologically active agents, as described herein. Any pharmaceuticallyacceptable biocompatible polymer known to those skilled in the art maybe utilized. It is preferred that the biocompatible controlled releasematerial degrade in vivo within about one year, preferably within about3 months, more preferably within about two months. More preferably, thecontrolled release material will degrade significantly within one tothree months, with at least 50% of the material degrading into non-toxicresidues, which are removed by the body, and 100% of the compound of theinvention being released within a time period within about two weeks,preferably within about 2 days to about 7 days. A degradable controlledrelease material should preferably degrade by hydrolysis, either bysurface erosion or bulk erosion, so that release is not only sustainedbut also provides desirable release rates. However, the pharmacokineticrelease profile of these formulations may be first order, zero order,bi- or multi-phasic, to provide the desired reversible localanti-nociceptive effect over the desired time period.

Suitable biocompatible polymers can be utilized as the controlledrelease material. The polymeric material may comprise biocompatible,biodegradable polymers, and, in certain preferred embodiments, ispreferably a copolymer of lactic and glycolic acid. Preferred controlledrelease materials which are useful in the formulations of the inventioninclude the polyanhydrides, polyesters, co-polymers of lactic acid andglycolic acid (preferably wherein the weight ratio of lactic acid toglycolic acid is no more than 4:1 i.e., 80% or less lactic acid to 20%or more glycolic acid by weight) and polyorthoesters containing acatalyst or degradation enhancing compound, for example, containing atleast 1% by weight anhydride catalyst such as maleic anhydride. Examplesof polyesters include polylactic acid, polyglycolic acid and polylacticacid-polyglycolic acid copolymers. Other useful polymers include proteinpolymers such as collagen, gelatin, fibrin and fibrinogen andpolysaccharides such as hyaluronic acid.

The polymeric material may be prepared by any method known to thoseskilled in the art. For example, where the polymeric material iscomprised of a copolymer of lactic and glycolic acid, this copolymer maybe prepared by the procedure set forth in U.S. Pat. No. 4,293,539,incorporated herein by reference. Alternatively, copolymers of lacticand glycolic acid may be prepared by any other procedure known to thoseskilled in the art. Other useful polymers include polylactides,polyglycolides, polyanhydrides, polyorthoesters, polycaprolactones,polyphosphazenes, polyphosphoesters, polysaccharides, proteinaceouspolymers, soluble derivatives of polysaccharides, soluble derivatives ofproteinaceous polymers, polypeptides, polyesters, and polyorthoesters ormixtures or blends of any of these.

Pharmaceutically acceptable polyanhydrides which are useful in thepresent invention have a water-labile anhydride linkage. The rate ofdrug release can be controlled by the particular polyanhydride polymerutilized and its molecular weight. The polysaccharides may bepoly-1,4-glucans, e.g., starch glycogen, amylose, amylopectin, andmixtures thereof. The biodegradable hydrophilic or hydrophobic polymermay be a water-soluble derivative of a poly-1,4-glucan, includinghydrolyzed amylopectin, derivatives of hydrolyzed amylopectin such ashydroxyethyl starch (HES), hydroxyethyl amylose, dialdehyde starch, andthe like. The polyanhydride polymer may be branched or linear.

Examples of polymers which are useful in the present invention include(in addition to homopolymers and copolymers of poly(lactic acid) and/orpoly(glycolic acid)) poly[bis(p-carboxyphenoxy) propane anhydride](PCPP), poly[bis(p-carboxy)methane anhydride](PCPM), polyanhydrides ofoligomerized unsaturated aliphatic acids, polyanhydride polymersprepared from amino acids which are modified to include an additionalcarboxylic acid, aromatic polyanhydride compositions, and co-polymers ofpolyanhydrides with other substances, such as fatty acid terminatedpolyanhydrides, e.g., polyanhydrides polymerized from monomers of dimersand/or trimers of unsaturated fatty acids or unsaturated aliphaticacids. Polyanhydrides may be prepared in accordance with the methods setforth in U.S. Pat. No. 4,757,128, incorporated herein by reference.Polyorthoester polymers may be prepared, e.g., as set forth in U.S. Pat.No. 4,070,347, incorporated herein by reference. Polyphosphoesters maybe prepared and used as set forth in U.S. Pat. Nos. 6,008,318,6,153,212, 5,952,451, 6,051,576, 6,103,255, 5,176,907 and 5,194,581,each of which is incorporated herein by reference.

Proteinaceous polymers may also be used. Proteinaceous polymers andtheir soluble derivatives include gelation biodegradable syntheticpolypeptides, elastin, alkylated collagen, alkylated elastin, and thelike. Biodegradable synthetic polypeptides includepoly-(N-hydroxyalkyl)-L-asparagine, poly-(N-hydroxyalkyl)-L-glutamine,copolymers of N-hydroxyalkyl-L-asparagine and N-hydroxyalkyl-L-glutaminewith other amino acids. Suggested amino acids include L-alanine,L-lysine, L-phenylalanine, L-valine, L-tyrosine, and the like.

In additional embodiments, the controlled release material, which ineffect acts as a carrier for a compound of the invention, alone or incombination with one or more biologically active agents as describedherein, can further include a bioadhesive polymer such as pectins(polygalacturonic acid), mucopolysaccharides (hyaluronic acid, mucin) ornon-toxic lectins or the polymer itself may be bioadhesive, e.g.,polyanhydride or polysaccharides such as chitosan.

In embodiments where the biodegradable polymer comprises a gel, one suchuseful polymer is a thermally gelling polymer, e.g., polyethylene oxide,polypropylene oxide (PEO-PPO) block copolymer such as Pluronic™ F127from BASF Wyandotte. In such cases, the local anesthetic formulation maybe injected via syringe as a free-flowing liquid, which gels rapidlyabove 30° C. (e.g., when injected into a patient). The gel system thenreleases a steady dose of a compound of the invention, alone or incombination with one or more biologically active agents as describedherein, at the site of administration.

Dosage Forms for Oral Use

Formulations for oral use include tablets containing the activeingredient(s) in a mixture with non-toxic pharmaceutically acceptableexcipients. These excipients may be, for example, inert diluents orfillers (e.g., sucrose, sorbitol, sugar, mannitol, microcrystallinecellulose, starches including potato starch, calcium carbonate, sodiumchloride, lactose, calcium phosphate, calcium sulfate, or sodiumphosphate); granulating and disintegrating agents (e.g., cellulosederivatives including microcrystalline cellulose, starches includingpotato starch, croscarmellose sodium, alginates, or alginic acid);binding agents (e.g., sucrose, glucose, sorbitol, acacia, alginic acid,sodium alginate, gelatin, starch, pregelatinized starch,microcrystalline cellulose, magnesium aluminum silicate,carboxymethylcellulose sodium, methylcellulose, hydroxypropylmethylcellulose, ethylcellulose, polyvinylpyrrolidone, or polyethyleneglycol); and lubricating agents, glidants, and antiadhesives (e.g.,magnesium stearate, zinc stearate, stearic acid, silicas, hydrogenatedvegetable oils, or talc). Other pharmaceutically acceptable excipientscan be colorants, flavoring agents, plasticizers, humectants, bufferingagents, taste masking agents (such as hydroxypropyl methylcellulose,hydroxypropyl cellulose), and the like.

One or more compounds of the invention and one or more biologicallyactive agents, as defined herein, may be mixed together in a tablet,capsule, or other vehicle, or may be partitioned. In one example, acompound of the invention is contained on the inside of the tablet, andthe biologically active agent is on the outside of the tablet, such thata substantial portion of the biologically active agent is released priorto the release of the compound of the invention.

Formulations for oral use may also be provided as chewable tablets, oras hard gelatin capsules wherein the active ingredient is mixed with aninert solid diluent (e.g., potato starch, lactose, microcrystallinecellulose, calcium carbonate, calcium phosphate or kaolin), or as softgelatin capsules wherein the active ingredient is mixed with water or anoil medium, for example, peanut oil, liquid paraffin, or olive oil.Powders, granulates, and pellets may be prepared using the ingredientsmentioned above under tablets and capsules in a conventional mannerusing, e.g., a mixer, a fluid bed apparatus or a spray drying equipment.

Formulations for oral administration to the mouth may also be providedas a mouthwash, an oral spray, oral rinse solution, or oral ointment ororal gel.

Dissolution or diffusion controlled release can be achieved byappropriate coating of a tablet, capsule, pellet, or granulateformulation of compounds, or by incorporating the compound into anappropriate matrix. A controlled release coating may include one or moreof the coating substances mentioned above and/or, e.g., shellac,beeswax, glycowax, castor wax, carnauba wax, stearyl alcohol, glycerylmonostearate, glyceryl distearate, glycerol palmitostearate,ethylcellulose, acrylic resins, dl-polylactic acid, cellulose acetatebutyrate, polyvinyl chloride, polyvinyl acetate, vinyl pyrrolidone,polyethylene, polymethacrylate, methylmethacrylate,2-hydroxymethacrylate, methacrylate hydrogels, 1,3 butylene glycol,ethylene glycol methacrylate, and/or polyethylene glycols. In acontrolled release matrix formulation, the matrix material may alsoinclude, e.g., hydrated methylcellulose, carnauba wax and stearylalcohol, carbopol 934, silicone, glyceryl tristearate, methylacrylate-methyl methacrylate, polyvinyl chloride, polyethylene, and/orhalogenated fluorocarbon.

The liquid forms in which the compounds and compositions of the presentinvention can be incorporated for administration orally include aqueoussolutions, suitably flavored syrups, aqueous or oil suspensions, andflavored emulsions with edible oils such as cottonseed oil, sesame oil,coconut oil, or peanut oil, as well as elixirs and similarpharmaceutical vehicles.

Generally, when administered to a human, the oral dosage of any of thecompounds of the combination of the invention will depend on the natureof the compound and can readily be determined by one skilled in the art.Typically, such dosage is normally about 0.001 mg to 2000 mg per day,desirably about 1 mg to 1000 mg per day, and more desirably about 5 mgto 500 mg per day. Dosages up to 200 mg per day may be necessary.

Administration of each drug in a combination therapy, as describedherein, can, independently, be one to four times daily for one day toone year, and may even be for the life of the patient. Chronic,long-term administration will be indicated in many cases.

Parenteral Formulations

Formulations suitable for parenteral administration (e.g., byinjection), include aqueous or non-aqueous, isotonic, pyrogen-free,sterile liquids (e.g., solutions, suspensions), in which the compound isdissolved, suspended, or otherwise provided (e.g., in a liposome orother microparticulate). Such liquids may additional contain otherpharmaceutically acceptable ingredients, such as antioxidants, buffers,preservatives, stabilizers, bacteriostats, suspending agents, thickeningagents, and solutes which render the formulation isotonic with the blood(or other relevant bodily fluid) of the intended recipient. Examples ofexcipients include, for example, water, alcohols, polyols, glycerol,vegetable oils, and the like. Examples of suitable isotonic carriers foruse in such formulations include Sodium Chloride Injection, Ringer'sSolution, or Lactated Ringer's Injection. Typically, the concentrationof the compound in the liquid is from about 1 ng/ml to about 10 μg/ml,for example from about 10 ng/ml to about 1 g/ml. The formulations may bepresented in unit-dose or multi-dose sealed containers, for example,ampoules and vials, and may be stored in a freeze-dried (lyophilized)condition requiring only the addition of the sterile liquid carrier, forexample water for injections, immediately prior to use. Extemporaneousinjection solutions and suspensions may be prepared from sterilepowders, granules, and tablets.

Formulations for Ophthalmic Administration

The compounds of the invention can also be formulated with anophthalmically acceptable carrier in sufficient concentration so as todeliver an effective amount of the active compound or compounds to theoptic nerve site of the eye. Preferably, the ophthalmic, therapeuticsolutions contain one or more of the active compounds in a concentrationrange of approximately 0.0001% to approximately 5% (weight by volume)and more preferably approximately 0.0005% to approximately 0.1% (weightby volume).

An ophthalmically acceptable carrier does not cause significantirritation to the eye and does not abrogate the pharmacological activityand properties of the charged sodium channel blockers.

Ophthalmically acceptable carriers are generally sterile, essentiallyfree of foreign particles, and generally have a pH in the range of 5-8.Preferably, the pH is as close to the pH of tear fluid (7.4) aspossible. Ophthalmically acceptable carriers are, for example, sterileisotonic solutions such as isotonic sodium chloride or boric acidsolutions. Such carriers are typically aqueous solutions contain sodiumchloride or boric acid. Also useful are phosphate buffered saline (PBS)solutions.

Various preservatives may be used in the ophthalmic preparation.Preferred preservatives include, but are not limited to, benzalkoniumpotassium, chlorobutanol, thimerosal, phenylmercuric acetate, andphenylmercuric nitrate. Likewise, various preferred vehicles may be usedin such ophthalmic preparation. These vehicles include, but are notlimited to, polyvinyl alcohol, povidone, hydroxypropyl methyl cellulose,poloxamers, carboxymethyl cellulose and hydroxyethyl cellulose.

Tonicity adjustors may be added as needed or convenient. They include,but are not limited to, salts, particularly sodium chloride, potassiumchloride, etc., mannitol and glycerin, or any other suitableophthalmically acceptable tonicity adjustor.

Various buffers and means for adjusting pH may be used so long as theresulting preparation is ophthalmically acceptable. Accordingly, buffersinclude but are not limited to, acetate buffers, citrate buffers,phosphate buffers, and borate buffers. Acids or bases may be used toadjust the pH of these formulations as needed. Ophthalmically acceptableantioxidants can also be include. Antioxidants include but are notlimited to sodium metabisulfite, sodium thiosulfate, acetylcysteine,butylated hydroxyanisole, and butylated hydroxytoluene.

Formulations for Nasal and Inhalation Administration

The pharmaceutical compositions of the invention can be formulated fornasal or intranasal administration. Formulations suitable for nasaladministration, when the carrier is a solid, include a coarse powderhaving a particle size, for example, in the range of approximately 20 to500 microns which is administered by rapid inhalation through the nasalpassage. When the carrier is a liquid, for example, a nasal spray or asnasal drops, one or more of the formulations can be admixed in anaqueous or oily solution and inhaled or sprayed into the nasal passage.

For administration by inhalation, the active ingredient can beconveniently delivered in the form of an aerosol spray presentation frompressurized packs or a nebulizer, with the use of a suitable propellant,e.g., dichlorodifluoromethane, trichlorofluoromethane,dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In thecase of a pressurized aerosol the dosage unit can be determined byproviding a valve to deliver a metered amount, Capsules and cartridgesof, for example, gelatin for use in an inhaler or insufflator can beformulated containing a powder mix of the compound and a suitable powderbase such as lactose or starch.

Dry powder compositions for topical delivery to the lung by inhalationmay, for example, be presented in capsules and cartridges of, forexample, gelatin or blisters of, for example, laminated aluminum foil,for use in an inhaler or insufflator. Powder blend formulationsgenerally contain a powder mix for inhalation of the compound of theinvention and a suitable powder base (carrier/diluent/excipientsubstance) such as mono-, di or ploy-saccharides (e.g. lactose orstarch). Use of lactose is preferred. In one embodiment, each capsule orcartridge may contain between about 2 ug to about 100 mg of the compoundof formula (I) optionally in combination with another therapeuticallyactive ingredient. In a preferred embodiment, each capsule or cartridgemay contain between about 10 ug to about 50 mg of the compound offormula (I) optionally in combination with another therapeuticallyactive ingredient. In another embodiment, each capsule or cartridge maycontain between about 20 ug to about 10 mg of the compound of formula(I) optionally in combination with another therapeutically activeingredient. Alternatively, the compound of the invention may bedelivered without excipients.

Suitably, the packaging/medicament dispenser is of a type selected fromthe group consisting of a reservoir dry powder inhaler (RDPI), amulti-dose dry powder inhaler (MDPI), and a metered dose inhaler (MDI).

Solutions or suspensions for use in a pressurized container, pump,spray, atomizer, or nebulizer can be formulated to contain an aqueousmedium, ethanol, aqueous ethanol, or a suitable alternative agent fordispersing, solubilizing, or extending release of the activeingredient(s); a propellant as solvent; and/or a surfactant, such assorbitan trioleate, oleic acid, or an oligolactic acid.

Compositions formulated for nasal or inhalation administration mayinclude one or more taste-masking agents such as flavoring agents,sweeteners, and other strategies, such as sucrose, dextrose, andlactose, carboxylic acids, menthol, amino acids or amino acidderivatives such as arginine, lysine, and monosodium glutamate, and/orsynthetic flavor oils and flavoring aromatics and/or natural oils,extracts from plants, leaves, flowers, fruits, etc. and combinationsthereof. These may include cinnamon oils, oil of wintergreen, peppermintoils, clover oil, bay oil, anise oil, eucalyptus, vanilla, citrus oilsuch as lemon oil, orange oil, grape and grapefruit oil, fruit essencesincluding apple, peach, pear, strawberry, raspberry, cherry, plum,pineapple, apricot, etc. Additional sweeteners include sucrose,dextrose, aspartame, acesulfame-K, sucralose and saccharin, organicacids (by non-limiting example citric acid and aspartic acid). Suchflavors may be present at from about 0.05 to about 4 percent by weight,and may be present at lower or higher amounts as a factor of one or moreof potency of the effect on flavor, solubility of the flavorant, effectsof the flavorant on solubility or other physicochemical orpharmacokinetic properties of other formulation components, or otherfactors.

Indications

The compounds, compositions, methods, and kits of the invention can beused to treat itch, pain, or cough. Conditions include trigeminaltrophic syndrome, erythromelalgia, back and neck pain, lower back pain,cancer pain, gynecological and labor pain, abdominal wall pain, chronicabdominal wall pain, fibromyalgia, allergic rhinitis, arthritis,rheumatoid arthritis, osteoarthritis, rheumatological pains, orthopedicpains, acute and post herpetic neuralgia and other neuropathic pains(including peripheral neuropathy), sickle cell crises, muscle pain,vulvodynia, rectal pain, Levator ani syndrome, proctalgia fugax,peri-anal pain, hemorrhoid pain, stomach pain, ulcers, inflammatorybowel disease, irritable bowel disease, irritable bowel syndrome, oralmucositis, esophagitis, interstitial cystitis, urethritis and otherurological pains, dental pain, burn pain, headaches, ophthalmicirritation, conjunctivitis (e.g., allergic conjunctivitis), eye redness,dry eye, dry eye syndrome (chronic ocular pain), complex regional painsyndrome, acute postoperative pain, postoperative pain, post-surgicalocular pain, and procedural pain (i.e., pain associated with injections,draining an abscess, surgery, dental procedures, ophthalmic procedures,ophthalmic irritation, conjunctivitis (e.g., allergic conjunctivitis),eye redness, dry eye, arthroscopies and use of other medicalinstrumentation, cosmetic surgical procedures, dermatologicalprocedures, setting fractures, biopsies, and the like).

Since a subclass of nociceptors mediate itch sensation, the compounds,compositions, methods, and kits of the invention are particularlysuitable to treat itch in patients with conditions like pruritus(including, but not limited to, brachioradial, chronic idiopathic,genital/anal, notalgia paresthetica, and scalp), allergic dermatitis,atopic dermatitis, contact dermatitis, poison ivy, infections,parasites, insect bites, pregnancy, metabolic disorders, liver or renalfailure, drug reactions, allergic reactions, eczema, hand eczema,genital and anal itch, hemorrhoid itch, and cancer.

Since a subclass of nociceptors can initiate aberrant cough reflexes,the compounds, compositions, methods, and kits of the invention can alsobe used to treat cough in patients with conditions like asthma, COPD,asthma-COPD overlap syndrome (ACOS), interstitial pulmonary fibrosis(IPF), idiopathic pulmonary fibrosis, post viral cough, post-infectioncough, chronic idiopathic cough and lung cancer.

The compounds, compositions, methods, and kits of the invention can alsobe used to treat neurogenic inflammation and neurogenic inflammatorydisorders. Inflammation is a complex set of responses to harmful stimulithat results in localized redness, swelling, and pain. Inflammation canbe innate or adaptive, the latter driven by antigens and is mediated byimmune cells (immune-mediated inflammation). Neurogenic inflammationresults from the efferent functions of pain-sensing neurons(nociceptors), wherein neuropeptides and other chemicals that arepro-inflammatory mediators are released from the peripheral terminals ofthe nociceptors when they are activated. This release process ismediated by calcium influx and exocytosis of peptide containingvesicles, and the pro-inflammatory neuropeptides include substance P,neurokinin A and B (collectively known as tachykinins), calcitoningene-related peptide (CGRP), and vasoactive intestinal polypeptide(VIP).

The release of peripheral terminal chemicals stimulate a variety ofinflammatory responses. First, the release of substance P can result inan increase in capillary permeability such that plasma proteins leakfrom the intravascular compartment into the extracellular space (plasmaextravasation), causing edema. This can be detected as a wheal (a firm,elevated swelling of the skin) which is one component of a triad ofinflammatory responses-wheal, red spot, and flare-known as the Lewistriple response. Second, the release of CGRP causes vasodilation,leading to increased blood flow. This can be detected as a flare, whichis another component of the Lewis triple response.

Substance P also has a pro-inflammatory action on immune cells (e.g.,macrophages, T-cells, mast cells, and dendritic cells) via theirneurokinin-1 (NK1) receptor. This effect has been documented in allergicrhinitis, gastritis, and colitis, and represents an interface betweenthe neurogenic and immune-mediated components of inflammation. SubstanceP released from one nociceptor may also act on NK1 receptors onneighboring nociceptors to sensitize or activate them, causing a spreadof activation and afferent/efferent function. These efferent functionsof nociceptors can be triggered by: 1) Direct activation of a nociceptorterminal by a peripheral adequate stimulus applied to the terminal (e.g.a pinch); 2) Indirect antidromic activation of a non-stimulatednociceptor terminal by the axon reflex, wherein action potential inputfrom one terminal of a nociceptor, upon reaching a converging axonalbranch point in the periphery, results in an action potential travelingfrom the branch point down to the peripheral terminal of anon-stimulated terminal; and 3) Activation as a result of activity innociceptor central terminals in the CNS traveling to the periphery(e.g., primary afferent depolarization of central terminals produced byGABA can be sufficient to initiate action potentials traveling the“wrong way”).

Genomic analysis of lung resident ILC2 cells has revealed expression ofreceptors for several neuropeptides released by sensory neurons,including SP, CGRP and VIP, providing an opportunity for nociceptors todirectly communicate with these cells. In particular, VIP is found to beexpressed in NaV1.8+ nodose ganglion neurons, including lung afferentsin OVA-exposed mice. Cultured nodose ganglion neurons stimulated withcapsaicin or IL5 also released VIP while BALF from OVA-exposed micecontained elevated VIP compared to vehicle-challenged mice (Talbot etal., Neuron. 15 Jul. 2015; 87(2): 341-354). These data indicate that VIPis released in the inflamed lung and can be blocked by silencing neuronswith charged sodium channel blockers of the present invention. Inaddition, when CD4+ T cells cultured under T_(H)2 skewing conditionswere exposed to recombinant mouse VIP, the transcript levels of IL-13and IL-5 increased, suggesting that VIP contributes to the competence ofT_(H)2 cells to transcribe these type II regulatory cytokines.

Nociceptors including those which are Nav1.8+/TRPV1⁺ have beendemonstrated to be integral in the immune response in models ofpsoriasis and contact dermatitis (Riol-Blanco et al., Nature Jun. 5,2014; 510(7503): 157-161). In the study on imiquimod-induced psoriasis,pharmacological or genetic ablation of nociceptors caused dermaldendritic cells (DDCs) to no longer no longer produce IL-23. This lackof IL-23 significantly reduced the production of inflammatory cytokinesdermal Th17 cells and also significantly reduced the influx ofinflammatory cells into the skin. By confocal microscopy 75% of DDCswere in either direct contact or in close proximity to sensory nerves.Pharmacological ablation of Nav1.8⁺/TRPV1⁺ nociceptors alsosignificantly reduced skin inflammation in the IL-12 driven DNFB modelof contact dermatitis (Riol-Blanco et al., Nature Jun. 5, 2014;510(7503): 157-161).

Immune mediator release from immune cells can also activate nociceptors.Mast cells are found close to primary nociceptive neurons and contributeto nociceptor sensitization in a number of contexts. Injection of thesecretagogue compound 48/80 promotes degranulation of mast cells in thedura and leads to excitation of meningeal nociceptors. Mast celldegranulation also contributes to the rapid onset of nerve growthfactor-induced thermal hyperalgesia. Macrophages contribute tonociceptor sensitization by releasing several soluble mediators.Expression of the chemokine macrophage inflammatory protein-1α (MIP-1α)and its receptors CCR1 and CCR5 is increased in macrophages and Schwanncells after partial ligation of the sciatic nerve and contributes to thedevelopment of neuropathic pain. Lymphocytes contribute to thesensitization of peripheral nociceptors. T cells infiltrate the sciaticnerve and dorsal root ganglion (DRG) after nerve injury. Hyperalgesiaand allodynia induced by nerve injury are markedly attenuated orabrogated in rodents lacking T cells and the immunosuppressant rapamycinattenuates neuropathic pain in rats, partly owing to an effect on Tcells. Among the subsets of T cells, type 1 and 2 helper T cells (T_(H)1and T_(H)2 cells) have been shown to have different roles in neuropathicpain. T_(H)1 cells facilitate neuropathic pain behavior by releasingproinflammatory cytokines (IL-2 and interferon-γ (IFNγ), whereas T_(H)2cells inhibit it by releasing anti-inflammatory cytokines (IL-4, IL-10and IL-13). The complement system also has a role in inflammatoryhyperalgesia and neuropathic pain. C5a, an anaphylatoxin, is animportant effector of the complement cascade and upon binding to C5aR1receptors on neutrophils it becomes a potent neutrophil attractant (Ren& Dubner, Nat. Med. 16:1267-1276 (2010)).

Bacterial infections have been shown to directly activate nociceptors,and that the immune response mediated through TLR2, MyD88, T cells, Bcells, and neutrophils and monocytes is not necessary for Staphylococcusaureus-induced pain in mice (Chiu et al., Nature 501:52-57 (2013)).Mechanical and thermal hyperalgesia in mice is correlated with livebacterial load rather than tissue swelling or immune activation.Bacteria induce calcium flux and action potentials in nociceptorneurons, in part via bacterial N-formylated peptides and thepore-forming toxin α-haemolysin, through distinct mechanisms. Specificablation of Nav1.8-lineage neurons, which include nociceptors, abrogatedpain during bacterial infection, but concurrently increased local immuneinfiltration and lymphadenopathy of the draining lymph node. Thus,bacterial pathogens produce pain by directly activating sensory neuronsthat modulate inflammation, an unsuspected role for the nervous systemin host-pathogen interactions. Data from Talbot et al., (Neuron. Jul.15, 2015; 87(2): 341-354) have also suggested that nociceptors areactivated during exposure to allergens in sensitized animals.

In certain disorders, neurogenic inflammation contributes to theperipheral inflammation elicited by tissue injury, autoimmune disease,infection, and exposure to irritants in soft tissue, skin, therespiratory system, joints, the urogenital and GI tract, the liver, andthe brain. Neurogenic inflammatory disorders include, but are notlimited to, allergic inflammation, inflammatory bowel disease,interstitial cystitis, atopic dermatitis, asthma, conjunctivitis,arthritis, colitis, contact dermatitis, diabetes, eczema, cystitis,gastritis, migraine headache, psoriasis, rhinitis, rosacea, sunburn,pancreatitis, chronic cough, chronic rhinosinusitis, traumatic braininjury, polymicrobial sepsis, tendinopathies, chronic urticaria,rheumatic disease, acute lung injury, exposure to irritants, inhalationof irritants, pollutants, or chemical warfare agents, as describedherein.

Assessment of Itch, Pain, Cough, and Neurogenic Inflammation

In order to measure the efficacy of any of the compounds, compositions,methods, and kits of the invention in the treatment of pain associatedwith musculoskeletal, immunoinflammatory and neuropathic disorders, ameasurement index may be used. Indices that are useful include a visualanalog scale (VAS), a Likert scale, categorical pain scales,descriptors, the Lequesne index, the WOMAC index, and the AUSCAN index,each of which is well known in the art. Such indices may be used tomeasure pain, itch, function, stiffness, or other variables.

A visual analog scale (VAS) provides a measure of a one-dimensionalquantity. A VAS generally utilizes a representation of distance, such asa picture of a line with hash marks drawn at regular distance intervals,e.g., ten 1-cm intervals. For example, a patient can be asked to rank asensation of pain or itch by choosing the spot on the line that bestcorresponds to the sensation of pain or itch, where one end of the linecorresponds to “no pain” (score of 0 cm) or “no itch” and the other endof the line corresponds to “unbearable pain” or “unbearable itch” (scoreof 10 cm). This procedure provides a simple and rapid approach toobtaining quantitative information about how the patient is experiencingpain or itch. VAS scales and their use are described, e.g., in U.S. Pat.Nos. 6,709,406 and 6,432,937.

A Likert scale similarly provides a measure of a one-dimensionalquantity. Generally, a Likert scale has discrete integer values rangingfrom a low value (e.g., 0, meaning no pain) to a high value (e.g., 7,meaning extreme pain). A patient experiencing pain is asked to choose anumber between the low value and the high value to represent the degreeof pain experienced. Likert scales and their use are described, e.g., inU.S. Pat. Nos. 6,623,040 and 6,766,319.

The Lequesne index and the Western Ontario and McMaster Universities(WOMAC) osteoarthritis index assess pain, function, and stiffness in theknee and hip of OA patients using self-administered questionnaires. Bothknee and hip are encompassed by the WOMAC, whereas there is one Lequesnequestionnaire for the knee and a separate one for the hip. Thesequestionnaires are useful because they contain more information contentin comparison with VAS or Likert. Both the WOMAC index and the Lequesneindex questionnaires have been extensively validated in OA, including insurgical settings (e.g., knee and hip arthroplasty). Their metriccharacteristics do not differ significantly.

The AUSCAN (Australian-Canadian hand arthritis) index employs a valid,reliable, and responsive patient self-reported questionnaire. In oneinstance, this questionnaire contains 15 questions within threedimensions (Pain, 5 questions; Stiffness, 1 question; and Physicalfunction, 9 questions). An AUSCAN index may utilize, e.g., a Likert or aVAS scale.

Indices that are useful in the methods, compositions, and kits of theinvention for the measurement of pain include the Pain Descriptor Scale(PDS), the Visual Analog Scale (VAS), the Verbal Descriptor Scales(VDS), the Numeric Pain Intensity Scale (NPIS), the Neuropathic PainScale (NPS), the Neuropathic Pain Symptom Inventory (NPSI), the PresentPain Inventory (PPI), the Geriatric Pain Measure (GPM), the McGill PainQuestionnaire (MPQ), mean pain intensity (Descriptor DifferentialScale), numeric pain scale (NPS) global evaluation score (GES) theShort-Form McGill Pain Questionnaire, the Minnesota MultiphasicPersonality Inventory, the Pain Profile and Multidimensional PainInventory, the Child Heath Questionnaire, and the Child AssessmentQuestionnaire.

Itch can be measured by subjective measures (VAS, Lickert, descriptors).Another approach is to measure scratch which is an objective correlateof itch using a vibration transducer or movement-sensitive meters.

Cough can be measured by standard questionnaires like the LeicesterCough Questionnaire as well as validated objective instruments tomeasure cough frequency (e.g., VitaloJAK).

EXAMPLES

The following examples are intended to illustrate the invention and arenot intended to limit it.

Example 1—Synthesis of Compounds 1A′ to 8A′ General AbbreviationDefinitions

-   ACN acetonitrile-   AcOH acetic acid-   aq. aqueous-   CDCl3 D₃-chloroform-   δ chemical shift (ppm)-   DCM dichloromethane-   DIPEA diisopropylethylamine-   DMSO dimethyl sulfoxide-   ESI electrospray ionization-   Et₂O diethyl ether-   EtOAc ethyl acetate-   h hour-   LAH lithium aluminum hydride-   MeOH methanol-   mHz megahertz-   MS mass spectrometry-   Ms mesylate (methane sulfonate)-   m/z mass to charge ratio-   NaCHBH₃ sodium cyanoborohydride-   NMR nuclear magnetic resounance-   pet ether petroleum ether-   RT room temperature-   TLC thin layer chromatography-   UV ultraviolet light

Synthesis of 1-benzyl-1-(2-(2, 6-dimethylphenoxy) ethyl) azepan-1-iumbromide

Synthesis of 2-(2-bromoethoxy)-1, 3-dimethylbenzene

To a stirred solution of 2, 6-dimethylphenol (0.50 g, 4.1 mmol) inacetonitrile (8.0 mL) was added potassium carbonate (1.7 g, 12.3 mmol)and 1, 2-dibromoethane (3.85 g, 20.5 mmol). The resulting reactionmixture was stirred at 90° C. for 16 h as progress of the reaction wasmonitored by TLC (Mobile phase: 5% EtOAc in pet ether, Visualization:UV). The reaction mixture was cooled to RT and concentrated underreduced pressure, diluted with water (100 mL) and extracted withdichloromethane (3×50 mL). The combined organic extracts were washedwith brine solution (1×50 ml), dried over anhydrous sodium sulphate andconcentrated under reduced pressure to afford crude which was purifiedby silica gel chromatography (eluted with pet ether) to afford pure2-(2-bromoethoxy)-1, 3-dimethylbenzene (0.20 g) as a colourless liquid.¹H NMR (400 MHz, CDCl₃) δ ppm 7.00-7.25 (m, 2H), 6.91-6.95 (m, 1H),4.01-4.10 (m, 2H), 3.65-3.68 (m, 2H), 2.30 (s, 6H).

Synthesis of intermediate 1-(2-(2, 6-dimethylphenoxy) ethyl) azepane

To a solution of 2-(2-bromoethoxy)-1, 3-dimethylbenzene (200 mg, 0.87mmol) in acetonitrile (3.0 mL) was added DIPEA (451.2 mg, 3.49 mmol) andazepane (120.2 mg, 1.22 mmol). The resulting reaction mixture wasstirred at 90° C. for 16 h as progress of the reaction was monitored byTLC. (Mobile phase: 10% EtOAc in pet ether, Visualization: UV). Thereaction mixture was cooled to RT and concentrated under reducedpressure, diluted with water (20 mL) and extracted with DCM (3×20 mL).The combined organic extracts were washed with brine solution (20 mL),dried over anhydrous sodium sulphate and concentrated under reducedpressure to afford 1-(2-(2, 6-dimethylphenoxy) ethyl) azepane (210 mg).¹H NMR (400 MHz, CDCl₃) δ ppm 6.98-7.00 (m, 2H), 6.88-6.92 (m, 1H),3.85-3.88 (m, 2H), 2.93-2.96 (m, 2H), 2.75-2.77 (m, 4H), 2.28 (s, 6H),1.59-1.68 (m, 8H).

Synthesis of 1-benzyl-1-(2-(2, 6-dimethylphenoxy) ethyl) azepan-1-iumbromide

To a solution of 1-(2-(2, 6-dimethylphenoxy) ethyl) azepane (100 mg, 0.4mmol) in acetonitrile (1.5 mL) was added benzyl bromide (0.072 ml, 0.6mmol) and the resulting reaction mixture was stirred at 90° C. for 16 hin sealed tube as progress of the reaction was monitored by TLC (Mobilephase: 50% EtOAc in pet ether, Visualization: UV). The reaction mixturewas cooled to room temperature and concentrated under reduced pressureto afford crude product which was triturated with ethyl acetate (3×5 ml)to afford 1-benzyl-1-(2-(2, 6-dimethylphenoxy) ethyl) azepan-1-iumbromide (80.5 mg) as an off white solid. MS (ESI): m/z 338.41 [M]+. ¹HNMR (400 MHz, DMSO-d₆) δ ppm 7.63-7.65 (m, 2H), 7.49-7.57 (m, 3H),7.04-7.08 (m, 2H), 6.96-6.98 (m, 1H), 4.72 (s, 2H), 4.29 (t, 2H),3.52-3.69 (m, 6H), 2.29 (s, 6H), 1.88-1.92 (m, 4H), 1.59-1.62 (m, 4H).

The following examples were synthesized according to proceduresdescribed for the compound 1A.

Com- MS pound (ESI): # Structure m/z 2A

310.2 [M] ⁺ 3A

324.2 [M] ⁺ 4A

352.3 [M] ⁺ 5A

366.3 [M] + 6A

380.3 [M] + 7A

394.3 [M] +

Synthesis of 1-benzyl-1-(2-(2,6-dimethylphenoxy)propyl)azocan-1-iumbromide

Example 2—Inhibition of Nav1.7 Current

The compounds were synthesized according to the described methods andtested for the ability to inhibit voltage-gated sodium channels.

Cell Culture

NaV1.7 was expressed upon induction with tetracycline. Cells werecultured in DMEM containing 10% dialyzed Fetal Bovine Serum (VWR,Radnor, Pa.), 1% Glutamax (VWR, Radnor, Pa.), 1% Penicillin-Streptomycin(VWR, Radnor, Pa.), 100 mg/L Hygromycin (Thermo Fisher Scientific,Waltham, Mass. and 5 mg/L Blasticidin (Alfa Aesar, Haverhill, Mass.).Cells were grown and maintained at 37° C. in a humidified environmentcontaining 10% C02 in air. Cells were detached from the culture flaskfor passage and harvested using 0.05% Trypsin-EDTA (Thermo FisherScientific, Waltham, Mass.). To induce NaV1.7, cells were induced withtetracycline (0.1-1 μg/mL, IBI Scientific, Peosta, Iowa) the day beforerecording and plated onto 24-well plates. Cells were washed with DPBS(VWR, Radnor, Pa.), trypsinized and then triturated five times in 10 mLof growth media to break apart cell aggregates. For one 24-well plate, 2mL of cell suspension was mixed with 23 mL of fresh growth media and0.1-1 μg/mL tetracycline added. 1 ml of mixed media with cells was thenadded to each well of a 24-well plate, with a 12 mm coverslip alreadyplaced in the bottom of the well. Cells were then incubated in 37° C.and 10% C02 overnight.

Patch Clamp Solutions & Drugs

The intracellular solution contained the following (in mM) CsCl 135,NaCl 10, EGTA 10, HEPES 10, MgCl₂ 2, adjusted to pH 7.2 with CsOH. Theexternal solution was a normal Ringer solution containing (in mM) NaCl155, HEPES 10, glucose 10, KCl 3.5, CaCl₂) 1.5, MgCl₂ 1 adjusted to pH7.4 with NaOH. CsCl is from Alfa Aesar, Haverhill, Mass. All otherchemicals are from Sigma-Aldrich, St. Louis, Mo. In order to test thedegree of internal block by test compounds the compounds were dissolvedin internal solution at the indicated test concentration. In controlexperiments the internal solution did not contain any compound. In orderto test the degree of external block by test compounds the compoundswere dissolved in external solution at the indicated test concentration.

Whole Cell Patch Clamp Protocol

18-24 hours after cells were induced with tetracycline, coverslips wereplaced into a chamber filled with Normal Ringer solution at roomtemperature and the chamber placed on a microscope. Pipettes were pulledfrom borosilicate glass on a P97 puller (Sutter Instrument, Novato,Calif.) and polished with a MF-830 Microforge (Narishige InternationalUSA, Inc, Amityville, N.Y.) to have a resistance of 1.5-2.5 MΩ whenfilled with CsCl internal solution at room temperature. Healthy cells(those that are round and translucent with no visible blemishes) werechosen for seal formation. A seal was formed between the pipette and thecell, and a brief pulse of suction was used to “break in” and establishthe whole-cell configuration. The membrane potential was held at −100 mVbefore the voltage protocol began. Only cells with series resistancebetween 1.5-5 MΩ were retained for analysis. The voltage protocol was asfollows: Cells were held at −100 mV for 12 ms followed by ahyperpolarizing step to −105 mV for 12 ms to monitor the leak. Cellswere then stepped back to −100 mV for 40 ms. Cells were then depolarizedto −20 mV for 10 ms and then returned to −100 mV for 26.

Internal Block by Test Compounds

Once the recording was started, the voltage protocol was run at 30second intervals for 5 minutes to get a stable baseline. This wasfollowed by four 30-second periods of 5 Hz stimulation of the samevoltage protocol separated by 1 minute of rest which was then followedby 0.33 Hz stimulation after the last train. Currents were recordedusing PatchMaster software with Heka EPC10 (HEKA Electronics, Lambrecht,Germany). Only cells with inward current amplitudes at −20 mV between400 pA and 4 nA were accepted. In addition, cells having leak currentsgreater than 10% of their current amplitudes were discarded.

Data Analysis: Internal Block

The data was plotted using the Patchmaster software (HEKA Electronics,Lambrecht, Germany) and analyzed by plotting the minimum current duringthe voltage step to −20 mV (peak inward current) as a function of time.In order to determine the degree of rundown over the course of anexperiment, the average peak inward current amplitude (2-3 points)before 5 Hz stimulation was designated as the baseline (I_(baseline)).The average peak inward current during the last 2 second of the last 5Hz train was measured (I_(test)). The control fraction current remainingwas calculated by dividing I_(test) by I_(baseline). On each recordingday three cells were tested with control internal solution and theaverage fraction of current remaining calculated (Ctrl fractioncurrent).

To determine the % block produced by test compounds applied internallythe following was done. The average peak inward current amplitude (2-3points) before 5 Hz stimulation was designated as 0% block(I_(0% block)). To correct for the current change under controlconditions, I_(0% block) was multiplied by the average Ctrl fractioncurrent remaining to get the corrected 0% block current. The averagepeak inward current during the last 2 seconds of the last 5 Hz train wasdesignated as the unblocked current (I_(unblocked)). The % block wascalculated using the following equation:(1−I_(unblocked)/(I_(0% block)*Ctrl fraction current remaining)×100).Representative compounds of Example 1 were tested for intracellularinhibition of NaV 1.7. Activity Range is % inhibition: “+++”>70%, “++”(70-40%) or “+” (<40%). The results are presented below.

Test Nav1.7 Extracellular Compound Concentration Inhibition 1A  1 μM +++2A 10 μM +++ 3A 10 μM +++ 4A  1 μM +++ 5A  3 μM ++ 6A  3 μM +++ 7A  3 μM+++ 8A  3 μM +++External Block by Test Compounds

Once the recording was started, the voltage protocol was run at 30second intervals for 5 minutes to get a stable baseline. This isfollowed by 5 Hz stimulation of the same voltage protocol run until theend of experiment. The test compound is added during the 5 Hzstimulation train making sure to wait until the cell shows stablecurrent rundown rate before addition of the compound. The test compoundis added for 5 minutes before washing out with normal Ringer's solution.Currents were recorded using PatchMaster software with Heka EPC10 (HEKAElectronics, Lambrecht, Germany). Only cells with inward currentamplitudes at −20 mV between 400 pA and 4 nA were accepted. In addition,cells having leak currents greater than 10% of their current amplitudeswere discarded.

Data Analysis: External Block

The data was plotted using the Patchmaster software (HEKA Electronics,Lambrecht, Germany) and analyzed by plotting the minimum current duringthe voltage step to −20 mV (peak inward current) as a function of time.To determine the % block produced by test compounds applied externallythe following was done. After the stable current rundown rate wasestablished during the 5 Hz stimulation train, the Rate_(rundown) wascalculated by dividing the change in peak current amplitude by time. Theaverage peak inward current amplitude (2-3 seconds) before addition ofcompound was used to determine 0% block (I_(0% block)). To correct forthe rundown, I_(0% block) is subtracted by the (Rate_(rundown)*5 min) toget the corrected 0% block current. The average peak inward currentduring the last 2-3 seconds of the 5 minutes of compound applicationtime before washing is the unblocked current (I_(unblocked)). The %block was then calculated using the following equation: Fraction currentblock=1−I_(unblocked)/(I_(0% block)−Rate_(rundown)*5 min).

Representative compounds of Example 1 were tested for extracellularinhibition of NaV 1.7. Activity Range is % inhibition: “+++”>70%, “++”(70-40%) or “+” (<40%). The results are presented below.

Test Nav1.7 Extracellular Compound Concentration Inhibition 1A 1 μM ++4A 1 μM ++

Example 3—Antipruritic Activity

The anti-pruritic activity of representative compounds of the inventionwas determined in C57BL/6 mice according to similar published procedures(Ramachandran et al, JPET 2020). Briefly, mice were anesthetized using2.5% isoflurane and a solution of test compound (10 mg/mL, 100 μL) wastopically applied to a shaven area at the nape using a solvent pipette(T=0). Test article was messaged into the skin until complete absorptionoccurred. Chloroquine (100 μg in 50 μL 0.9% saline) was injectedintradermal at the indicated time to induce itch. Scratching activitywas monitored over 40 minutes.

Representative compounds of Example 1 significantly reducechloroquine-induced itch up to 8 hours following topical administration.See FIGS. 1A, 1B and 1C.

The patent and scientific literature referred to herein establishes theknowledge that is available to those with skill in the art. All UnitedStates patents and published or unpublished United States patentapplications cited herein are incorporated by reference. All publishedforeign patents and patent applications cited herein are herebyincorporated by reference. All other published references, documents,manuscripts and scientific literature cited herein are herebyincorporated by reference.

While this invention has been particularly shown and described withreferences to preferred embodiments thereof, it will be understood bythose skilled in the art that various changes in form and details may bemade therein without departing from the scope of the inventionencompassed by the appended claims. It will also be understood that noneof the embodiments described herein are mutually exclusive and may becombined in various ways without departing from the scope of theinvention encompassed by the appended claims.

What is claimed is:
 1. A compound represented by Formula (I):

wherein: Y⁻ is a pharmaceutically acceptable anion; wherein R₁, R₂, R₃,R₄, R₅, and R₆ are independently selected from hydrogen, C1-C4 alkyl,C1-C4 heteroalkyl, aryl or heteroaryl n is 0, 1, 2, 3, 4 and 5; q is 0,1, 2, 3, 4 or
 5. 2. The compound of claim 1, wherein R₁, R₂, R₃, R₄, R₅,and R₆ are hydrogen.
 3. The compound of claim 1, wherein q is
 2. 4. Thecompound of claim 3, wherein R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen. 5.The compound of claim 1, wherein q is
 1. 6. The compound of claim 5,wherein R₁, R₂, R₃, R₄, R₅, and R₆ are hydrogen.
 7. The compound ofclaim 1, wherein q is
 0. 8. The compound of claim 7, wherein R₁, R₂, R₃,R₄, R₅, and R₆ are hydrogen.
 9. The compound of claim 1, wherein R₁ ismethyl R₂, R₃, R₄, R₅, and R₆ are hydrogen.
 10. The compound of claim 1,wherein R₃ is methyl R₁, R₂, R₄, R₅, and R₆ are hydrogen.
 11. Thecompound of claim 1, wherein n is
 0. 12. The compound of claim 1,wherein n is
 1. 13. The compound of claim 1, wherein n is
 2. 14. Thecompound of claim 1, wherein n is
 3. 15. The compound of claim 1,wherein n is
 4. 16. The compound of claim 1, wherein n is
 5. 17. Thecompound of claim 1, wherein Y− is bromide or chloride.
 18. Apharmaceutical composition comprising the compound of claim 1 and apharmaceutically acceptable excipient.
 19. The composition of claim 18,wherein said composition is formulated for topical or dermaladministration.
 20. A method for treating itch, pain, cough, or aneurogenic inflammatory disorder in a patient, comprising administeringto said patient an effective amount of the compound of claim
 1. 21. Themethod of claim 20, wherein said itch is selected from the groupconsisting of itch due to pruritus, brachioradial pruritus, chronicidiopathic pruritus, genital/anal pruritus, notalgia paresthetica, scalppruritus, allergic dermatitis, contact dermatitis, atopic dermatitis,psoriasis, prurigo nodularis, hand eczema, poison ivy, sunburn, dryskin, infections, parasites, insect bites, pregnancy, metabolicdisorders, liver or renal failure, drug reactions, allergic reactions,eczema, genital and anal itch, hemorrhoid itch, and cancer.